Project Summary The overall goal of our research program is to elucidate the underlying molecular principles that govern synaptic plasticity associated with chronic pain. Chronic neuropathic pain is a significant and unmet clinical problem. Glutamate ?-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs) mediate the vast majority of fast excitatory synaptic transmission in the mammalian central nervous system. AMPARs are tetrameric cation channels composed of a combinational assembly of four subunits, GluA1 through GluA4. GluA2 is particularly important for the biophysical properties of AMPARs because GluA2-containing AMPARs are impermeable to Ca2+. In contrast, GluA2-lacking AMPARs show inward-rectifying currents and have a high Ca2+ permeability and are thus referred to as Ca2+-permeable AMPARs (CP-AMPARs). The prevalence of synaptic CP-AMPARs of spinal dorsal horn neurons is markedly increased in neuropathic pain. However, the molecular mechanisms underlying the switch of AMPAR subunit composition in neuropathic pain remain little known. The major objective of our proposal is to determine the key molecular mechanism responsible for regulating the assembly and trafficking of CP-AMPARs in neuropathic pain. ?2?-1, often considered a Ca2+ channel subunit, is upregulated in the spinal dorsal horn in neuropathic pain. Our preliminary studies showed that ?2?-1 interacted with AMPAR subunits in vitro and in vivo and that increased ?2?-1 expression promoted synaptic incorporation of CP-AMPARs in the spinal dorsal horn. In this proposal, we will test our overall hypothesis that ?2?-1 potentiates the synaptic CP-AMPAR prevalence in spinal dorsal horn neurons in neuropathic pain through physical interaction with AMPAR subunits to preferentially regulate their subunit composition and synaptic trafficking. We will use a multidisciplinary approach to study ?2?-1?AMPAR coupling and its distinct role in neuropathic pain at molecular, cellular, and behavioral levels. At the completion of our project, we will gain significant mechanistic insight into the poorly defined role of ?2?-1 in synaptic plasticity and neuropathic pain caused by nerve injury and diabetic neuropathy. This new information will redefine the physiological function ?2?-1 and the role of ?2?-1?bound CP-AMPARs in the therapeutic effects of gabapentinoids. Therefore, the findings from the proposed studies will have a sustained positive impact by advancing our understanding of the synaptic mechanism of neuropathic pain, leading to the development of new therapies for chronic neuropathic pain.